2.1. The T Cell Antigen Receptor
T lymphocytes interact with antigens through the T cell antigen receptor (TCR) complex. The TCR is a clone-specific heterodimer on T cells, which recognizes its target antigen in association with a major histocompatiblity antigen. The TCR has been shown to be noncovalently associated with the CD3 complex. TCR is highly polymorphic between T cells of different specificities. Approximately 90 percent of peripheral blood T cells express a TCR consisting of an .alpha. polypeptide and a .beta. polypeptide. A small percentage of T cells have been shown to express a TCR consisting of a .gamma. polypeptide and a .delta. polypeptide. (Regarding TCR molecules, see Davis and Bjorkman, 1988, Nature 334:395-402; Marrack and Kappler, 1986, Sci. Amer. 254: 36; Meuer et al., 1984, Ann. Rev. Immunol. 2:23-50; Brenner et al., 1986, Nature 322:145-159; Krangel et al., 1987, Science 237:1051-1055; Hata et al., 1987, Science 238:678-682; Hochstenbach et al., 1988, J. Exp. Med. 168:761-776). The chains of the T cell antigen receptor of a T cell clone are each composed of a unique combination of domains designated variable (V), [diversity (D),] joining (J), and constant (C) (Siu et al., 1984, Cell 37:393; Yanagi et al., 1985, Proc. Natl. Acad. Sci. U.S.A. 82:3430). Hypervariable regions have been identified (Patten et al., 1984, Nature 312:40; Becker et al., 1985, Nature 317:430). In each T cell clone, the combination of V, D and J domains of both the alpha and the beta chains or of both the delta and gamma chains participates in antigen recognition in a manner which is uniquely characteristic of that T cell clone and defines a unique binding site, also known as the idiotype of the T cell clone. In contrast, the C domain does not participate in antigen binding.
2.2. T Cell Antigen Receptor Genes
TCR genes, like immunoglobulin genes, consist of regions which rearrange during T cell ontogeny (Chien et al., 1984, Nature 312:31-35; Hedrick et al., 1984, Nature 308:149-153; Yanagi et al., 1984, Nature 308:145-149). In genomic DNA, each TCR gene has V, J, and C regions; TCR .beta. and .delta. polypeptides also have D regions. The V (variable), D (diversity), J (junctional) and C (constant) regions are separated from one another by spacer regions in the DNA. There are usually many variable region segments and somewhat fewer diversity, junctional, and constant region segments. As a lymphocyte matures, these various segments are spliced together to create a continuous gene sequence consisting of one V, (D), J, and C region. TCR diversity, and thereby T cell specificity, is derived from several sources (Barth et al., 1985, Nature 316:517-523; Fink et al., 1986, Nature 321:219-225): a multiplicity of germline gene segments (Chien et al., 1984, Nature 309:322-326; Malissen et al., 1984, Cell 37:1101-1110; Gascoigne et al., 1984, Nature 310:387-391; Kavaler et al., 1984, Nature 310: 421-423; Siu et al., 1984, Nature 311:344-349; Patten et al., 1984, Nature 312:40-46), combinatorial diversity through the assembly of different V, D, J, and C segments (Siu et al., 1984, Cell 37:393-401; Goverman et al., 1985, Cell 40:859-867), and junctional flexibility, N-region diversity and the use of either multiple D regions or any of the three translational reading frames for D.beta. segments. TCR diversity does not appear to arise from the somatic hypermutation mechanism observed for immunoglobulins (Barth et al., supra). As a result of these mechanisms, TCRs are generated which differ in their amino-terminal, or N-terminal, domains (called variable, or V regions, constructed from combinations of V, D, and J gene segments) but are the same elsewhere, including their carboxy-terminal, or C-terminal domains (called constant regions). Accordingly, an almost limitless repertoire of TCR is established.
The V.beta. gene of the TCR appears to resemble most closely the immunoglobulin V gene in that it has three gene segments, V.beta., D.beta., and J.beta., which rearrange to form a contiguous V.beta. gene (Siu et al., 1984, Cell 37:393-401). The .beta. locus has been well characterized in mice, where it spans 700-800 kilobases of DNA and is comprised of two nearly identical C regions tandemly arranged with one D element and a cluster of 5-6 J elements 5' to each (Kronenberg et al., 1986, Ann. Rev. Immunol. 3:537-560). Approximately twenty to thirty V.beta. regions are located upstream (5') to the D, J, and C elements (Behlke et al., 1985, Science 229:566-570) although V.beta. genes may also be located 3' to the murine C.beta. genes (Malissen et al., 1986, Nature 319:28). Study of the structure and diversity of the human TCR .beta.-chain variable region genes has led to the grouping of genes into distinct V.beta. subfamilies (Tillinghast et al., 1986, Science 233:879-883; Concannon et al., 1986, Proc. Natl. Acad. Sci. U.S.A. 83:6598-6602; Borst et al., 1987, J. Immunol. 139:1952-1959).
The .gamma.-TCR gene was identified, first in mice (Saito et al., 1984, Nature 309:757-762; Kranz et al., 1985, Nature 313:762-755; Hayday et al., 1985, Cell 40:259-269) and then in humans (Lefranc et al., 1985, Nature 316:464-466; Murre et al., 1985, Nature 316:549-552). The human .gamma.TCR locus appears to consist of between five and ten variable, five joining, and two constant region genes (Dialynas et al., 1986, Proc. Natl. Acad. Sci. U.S.A. 83:2619).
The TCR .alpha. and .delta. locus are next to one another on human chromosome 14. Many TCR .delta. coding segments are located entirely within the .delta. gene locus (Satyanarayana et al., 1988, Proc. Natl. Acad. Sci. U.S.A. 85:8166-8170 Chien et al., 1987, Nature 330:722-727; Elliot et al., 1988, Nature 331:627-631). It is estimated that there are a minimum of 45-50 V.alpha. regions (Becker et al., Nature 317:430-434) whereas there are only approximately 10 V.delta. regions (Chien et al., 1987, supra). In peripheral blood, two predominant V.delta. genes appear to be expressed, namely, V.delta.1 and V.delta.2, identifiable by monoclonal antibodies, .delta.TCS1 and BB3, respectively. Nucleic acid sequences of TCR a genes have been reported (Sim et al., 1984, Nature 312:771-775; Yanagi et al., 1985, Proc. Natl. Acad. Sci. USA 82:3430-3434; Berkout et al., 1988, Nucl. Acids Res. 16:5208).
2.3. Antibodies to the T Cell Antigen Receptor
Clonotypic antibodies react only with a particular clone of T cells. Acuto et al. produced clonotypic monoclonal antibodies against a human thymocyte cell line, and thereby identified the TCR in relatively undifferentiated T3.sup.+ cells (1983, Cell 34:717-726). Meuer et al. showed that anti-TCR clonotypic monoclonal antibodies coupled to sepharose beads could induce production of interleukin-2 (1984, Proc. Natl. Acad. Sci. 81:1509-1513). Anti-TCR clonotypic antibody directed toward the CT8 cell line could only block cytotoxic effector cell function of that T cell line (Meuer et al., 1984, Ann. Rev. Immunol. 2:23-50). Antibodies which recognize TCR from many T cell lines recognize shared epitopes, or framework regions, of TCR peptides. Brenner et al. found that different cloned T cell lines shared antigenic determinants, none of which appeared to be accessible at the cell surface (1984, J. Exp. Med. 160:541-551). .beta.-Framework-1 (.beta.F1) monoclonal antibody reacts with a "hidden determinant" on the surface of viable T cells, and recognizes the TCR .beta. polypeptide in Western blots (Brenner et al., 1987, J. Immunol. 138:1502-1509). Another framework antibody, WT31, originally thought to be a framework reagent is useful in cell binding, but is inefficient in immunoprecipitation studies (Spits et al., 1985, J. Immunol. 135:1922-1928). WT31 now appears to recognize a CD3 determinant.
2.4. Rheumatoid Arthritis
Rheumatoid arthritis (RA), a chronic, recurrent, inflammatory disease primarily involving joints, affects 1-3% of North Americans, with a female to male ratio of 3:1. Severe RA patients tend to exhibit extra-articular manifestations including vasculitis, muscle atrophy, subcutaneous nodules, lymphadenopathy, splenomegaly and leukopenia. Spontaneous remission may occur; other patients have brief episodes of acute arthritis with longer periods of low-grade activity; still others progress to severe deformity of joints. In some patients with rheumatoid arthritis, particularly those with long-standing disease, a constellation of symptoms called "Felty's syndrome" develops, in which the typical arthropathy is accompanied by splenomegaly and neutropenia. It is estimated that about 15% of RA patients (severe RA and Felty's syndrome) become completely incapacitated ("Primer on the Rheumatic Diseases, 8th edition, 1983, Rodman, G. P. & Schumacher, H. R., Eds., Zvaifler, N. J., Assoc. Ed., Arthritis Foundations, Atlanta, Ga.).
The antigenic stimulus initiating the immune response and consequent inflammation is unknown. Certain HLA types (DR4, Dw4, Dw14 and DR1) have an increased prevalence of RA, perhaps leading to a genetic susceptibility to an unidentified factor which initiates the disease process. The association with DR4 is highest for Felty's Disease and severe RA (Westedt, M. L., et al., Annals of Rheumatic Diseases, 1986, 45, 534-538). Relationships between Epstein Barr virus and RA have been suggested. Synovial lymphocytes produce IgG that is recognized as foreign and stimulates a local immune response with production of anti-IgG-antibodies (rheumatoid factors). Immune complexes are formed by activation of the complement system which results in inflammation including activation of lysozyme and other enzymes. Helper T cell infiltration of the synovium and liberation of lymphokines such as IL6 lead to further accumulation of macrophages and slowly progressing joint destruction (erosions).
The approach to drug treatment in rheumatoid arthritis has been described as a pyramid ("Primer on the Rheumatic Diseases", supra). First line agents include aspirin and NSAIDS (non-steroidal anti-inflammatory drugs). When these agents fail, gold salts, penicillamine, methotrexate, or antimalarials, known as conventional second line drugs, are considered. Finally, steroids or cytotoxics are tried in patients with serious active disease that is refractory to first and second line treatment. Cyclosporine is now suggested to have a role in the treatment of patients whose disease is unresponsive to aspirin, NSAIDS, gold or penicillamine. However, the current experimental drugs to treat severe RA patients may prove too toxic even if they are effective.
Numerous efforts have been directed to developing safer and more efficacious immunotherapy to replace these toxic drugs. Severe RA patients who were treated with total lymphoid irradiation or thoraic duct drainage experienced significant improvement of disease symptoms. These procedures are not suitable for routine application. Due to these encouraging findings, however, and to the demonstration of the presence of T cells in the synovial infiltrate, it is possible to design new immunotherapies to specifically eliminate T cells. Most of these new experimental immunotherapies are targeted toward all or the bulk of T cells, and thus may produce significant side effects. A better approach for selective immunotherapy may be to eliminate only the small proportion of T cells that are involved in RA.
2.5. Role of T Cells in Rheumatoid Arthritis
Evidence has accumulated supporting a role for T-cells in the pathogenesis of rheumatoid arthritis (RA). The synovial tissue and surrounding synovial fluid of patients with rheumatoid arthritis (RA) are infiltrated with large numbers of cells. Activated and resting T cells can mediate tissue damage by a variety of mechanisms including the direct cytotoxicity of target cells expressing specific antigen in combination with the appropriate HLA restricting elements. The strong association of certain HLA products with RA has led researchers to implicate T cells in the autoimmune destruction of RA patient joints. In fact, HLA DR4, Dw4 and Dw14 gene products are among the major class II molecules that contribute significantly to disease susceptibility in RA patients (Nepom, B., et al., 1987, Abstracts of Amer. Rheumatism Assoc., p. S25; Todd, J. A., et al., 1988, Science 240:1003-1009), and they are capable of restricting antigen recognition of CD4+ T cells, primarily. Other autoimmune diseases also show a high correlation between disease susceptibility and HLA expression (Table 1).
This genetic basis of disease risk has resulted in phenotypic analysis of the T cells found within diseased joints. Previously, comparisons of T cells from RA joints and RA peripheral blood (PB) demonstrated significant differences in CD4 or CD8 phenotype, therefore implying a selection of T cells involved in disease activity. Most studies agree that synovial tissue-infiltrated T cells were mostly CD4+ helper-inducer (4B4+) cells (Duke, O., et al., 1987, Arth. Rheum., 30, 849) while the PB usually contained a mixture of CD4+ and CD8+ cells including both helper-inducer cells and suppressor-inducer cells (2H4+) (Emery, P., et al., 1987, Arth. Rheum., 30, 849). In contrast, there is additional evidence that the CD4+ infiltrate may be predominantly suppressor-inducer cells (2H4+) (Mikasaka, N., et al., 1987, Amer. Rheum. Abstracts, p. S39).
2.6. .gamma..delta.Positive T Cells
.gamma..delta. TCR may be the principal TCR in selected sites such as the skin or other organs. Although the function of the .gamma..delta. positive T cells is largely unknown, they appear to be involved in non-MHC-restricted cytotoxicity and IFN-.gamma. production. .gamma..delta..sup.+ T cells are known to secrete a variety of lymphokines, such as TNF alpha, IL2 and IL4 (Bluestone, J. A. and Matis, L. A., 1989, J. Immunol. 142, 1785-1788). The total population of .delta.+ T cells can be identified by the monoclonal antibody, TCR.delta.1, which recognizes a major framework determinant on the .delta. TCR (Band, H., et al., 1987, Science, 238, 682). A subset of .gamma..delta. positive T-lymphocytes can be identified by the monoclonal antibodies, .delta.TCS1 (anti-V.sub..delta. 1; Wu, Y-J., et al., 1988, J. Immunol. 141, 1476-1479) and BB3 (anti-V.sub..delta. TCR, Bottino, C., et al., 1988, J. Exp. Med., .about..about., 491-505). A study by Grossi, C. E., et al. (Proc. Natl. Acad. Sci., 1989) indicated that .delta.TCS1.sup.+ T cells exhibit motile cell morphology and migrate in tissue culture. .delta.TCS1.sup.+ T cells were also shown to be potent killer T cells (Rivas, A., et al., 1989, J. Immunol., 142, 1840-1846).